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Principal Investigator: Beth A Shen
Organization: UNIVERSITY OF WASHINGTON
Fiscal Year: 2024
Award: $78,892
Funding agency: National Institute of Allergy and Infectious Diseases
Abstract
Decades of research have revealed that the composition of the human gut microbiome directly impacts human
health, yet we still do not understand the mechanisms underlying formation and maintenance of this complex
community. Counterintuitively, increasing evidence suggests that competitive interactions play a role in
maintaining community stability. While genes encoding components of diverse systems involved in interbacterial
antagonism are widespread in human gut metagenomes, the specific antagonistic interactions occuring in situ
remain elusive. One of the pathways mediating interbacterial antagonism is the Esx secretion system, which is
conserved in two prominent gut phyla, Firmicutes and Actinobacteria. While this pathway has been shown to
mediate contact-dependent interbacterial antagonism by a limited number of Gram-positive species in vitro,
many questions remain about its physiological function. Antibacterial toxins secreted by the Esx machinery are
present throughout human gut metagenomes, suggesting that Esx-mediated antagonism could occur in this
community. These findings led me to propose to identify the targets of Esx-mediated interbacterial
antagonism in a model natural community, the murine gut microbiome. To accomplish this goal, I will take
advantage of the fact that bacteria express cognate immunity genes to prevent self-intoxication by secreted
toxins. In Aim 1 of my proposed studies, I will use bioinformatics and in vitro functional assays to identify Esx
toxin and immunity genes encoded in a model murine gut metagenome derived from wild-caught mice. My
preliminary findings indicate at least two antibacterial Esx toxins are encoded in this natural gut microbiome. In
Aim 2, I describe an in situ conjugation strategy to deliver the immunity genes identified in Aim 1 to all members
of the murine gut community. I hypothesize that expression of immunity genes in bacteria targeted by specific
Esx toxins will increase their abundance in a community. Therefore, by identifying changes to community
composition that depend on immunity genes, I can determine the targets of Esx-mediated antagonism in situ.
This unique approach to examine interbacterial antagonism under physiological conditions will significantly
improve our understanding of the physiological function of the Esx system in gut bacterial species. It will also
provide the first direct characterization of antagonism between gut microbiome constituents. The methods I
propose to develop may additionally later be applied broadly to the characterization of diverse antagonistic
pathways present in gut bacterial species, an essential next step needed to define the mechanisms shaping gut
microbiome composition and ultimately human health. By working in the lab of Dr. Joseph Mougous at the
University of Washington and with my experienced collaborators, I will be able to learn diverse techniques,
practice effective mentoring strategies, and hone my scientific communication skills. This high-quality training
will enable me to succeed as an independent researcher studying microbe-microbe interactions.
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